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SeqShop: Sequence Mapping and Assembly Practical, June 2014 (view source)

Revision as of 11:07, 2 February 2017

871 bytes added ,  11:07, 2 February 2017
→‎Download the example data
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'''Note:''' the latest version of this practical is available at: [[SeqShop: Sequence Mapping and Assembly Practical]]
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* The ones here is the original one from the June workshop (updated to be run from elsewhere)
 +
 
== Introduction ==
 
== Introduction ==
 
See the [[Media:SeqShop - GotCloud Align.pdf|introductory slides]] for an intro to this tutorial.
 
See the [[Media:SeqShop - GotCloud Align.pdf|introductory slides]] for an intro to this tutorial.
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''If you are running during the SeqShop Workshop, please skip this section.''
 
''If you are running during the SeqShop Workshop, please skip this section.''
 
<div class="mw-collapsible-content">
 
<div class="mw-collapsible-content">
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=== Download the example data ===
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=== Download & Build GotCloud ===
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If you do not already have GotCloud:
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* cd to where you want GotCloud installed (you can change this to any directory you want)
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mkdir -p ~/seqshop
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cd ~/seqshop/
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* download, decompress, and build the version of gotcloud that was tested with this tutorial:
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wget https://github.com/statgen/gotcloud/archive/gotcloud.workshop.tar.gz
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tar xvf gotcloud.workshop.tar.gz
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mv gotcloud-gotcloud.workshop gotcloud
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cd gotcloud/src
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make
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cd ../..
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=== Setup your run environment ===
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Remember the path to gotcloud/ that is what you will need to set your GC variable to.
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Environment variables will be used throughout the tutorial.
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=== Download the example data ===
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Download and untar file containing the example data used in the practicals:
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We recommend that you setup these variables so you won't have to modify every command in the tutorial.
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wget http://csg.sph.umich.edu//mktrost/seqshopExample.tar.gz
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tar xvf seqshopExample.tar.gz
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<div class="mw-collapsible mw-collapsed" style="width:500px">
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I'm using bash (replace the paths below with the appropriate paths):
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<div class="mw-collapsible-content">
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* Point to where you installed GotCloud
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*:<pre>export GC=/home/username/gotcloud</pre>
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* Point to where you installed the seqshop files
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*:<pre>export SS=/home/username/seqshop/</pre>
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* Point to where you want the output to go
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*:<pre>export OUT=/home/username/seqshop_output/</pre>
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</div>
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</div>
     −
<div class="mw-collapsible mw-collapsed" style="width:500px">
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You will see the names of all the files included in the example data scrolling on the screen as they are unpacked from the tar file.
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I'm using tcsh (replace the paths below with the appropriate paths):
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<div class="mw-collapsible-content">
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* Point to where you installed GotCloud
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*:<pre>setenv GC /home/username/gotcloud</pre>
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* Point to where you installed the seqshop files
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*:<pre>setenv SS /home/username/seqshop/</pre>
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* Point to where you want the output to go
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*:<pre>setenv OUT /home/username/seqshop_output/</pre>
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</div>
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</div>
     −
</div>
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{{SeqShopRemoteEnv}}
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</div>
      
== Examining [[GotCloud]] Align Input Files ==
 
== Examining [[GotCloud]] Align Input Files ==
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<li>View Screenshot</li>
 
<li>View Screenshot</li>
 
<div class="mw-collapsible-content">
 
<div class="mw-collapsible-content">
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[[File:RefDir.png|500px]]
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[[File:RefDir.png|700px]]
 
</div>
 
</div>
 
</div>
 
</div>
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Remember, use <code>'q'</code> to exit out of <code>less</code>
 
Remember, use <code>'q'</code> to exit out of <code>less</code>
 
  q
 
  q
−
  −
If you want to access the FASTA file by position, you can use <code>samtools faidx</code> command
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${GC}/bin/samtools faidx ${SS}/ref22/human.g1k.v37.chr22.fa 22:36000000 | less
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or
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${GC}/bin/samtools faidx ${SS}/ref22/human.g1k.v37.chr22.fa 22:36000000-36000100
      
; Where is the reference sequence?
 
; Where is the reference sequence?
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</ul>
 
</ul>
 
</ul>
 
</ul>
 +
 +
If you want to access the FASTA file by position, you can use <code>samtools faidx</code> command
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${GC}/bin/samtools faidx ${SS}/ref22/human.g1k.v37.chr22.fa 22:36000000 | less
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or
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${GC}/bin/samtools faidx ${SS}/ref22/human.g1k.v37.chr22.fa 22:36000000-36000100
    
=== GotCloud FASTQ Index File ===
 
=== GotCloud FASTQ Index File ===
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Let's look a look at the index file I prepared for this tutorial:
 
Let's look a look at the index file I prepared for this tutorial:
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  less -S ${IN}/align.index  
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  less -S ${SS}/align.index  
    
Remember, use <code>'q'</code> to exit out of <code>less</code>
 
Remember, use <code>'q'</code> to exit out of <code>less</code>
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<li>Need a reminder of the format?</li>
 
<li>Need a reminder of the format?</li>
 
<div class="mw-collapsible-content">
 
<div class="mw-collapsible-content">
−
[[File:fqindex.png|650px]]
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[[File:fqindex.png|750px]]
 
</div>
 
</div>
 
</div>
 
</div>
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<ul>
 
<ul>
 
<li>Use cut to extract just the MERGE_NAME & RGID fields </li>
 
<li>Use cut to extract just the MERGE_NAME & RGID fields </li>
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  cut -f 1,4 ${IN}/align.index
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  cut -f 1,4 ${SS}/align.index
 
</ul>
 
</ul>
 
</div>
 
</div>
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<li>HG00553 & HG00640</li>
 
<li>HG00553 & HG00640</li>
 
<li>They have multiple unique values in the RGID field</li>
 
<li>They have multiple unique values in the RGID field</li>
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[[File:fqindexRG.png|650px]]
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[[File:fqindexRG.png|800px]]
 
</div>
 
</div>
 
</div>
 
</div>
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Let's look at the configuration file I created for this test:
 
Let's look at the configuration file I created for this test:
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  more ${IN}/gotcloud.conf
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  more ${SS}/gotcloud.conf
    
Use the <code>space bar</code> to advance if the whole file isn't displayed.
 
Use the <code>space bar</code> to advance if the whole file isn't displayed.
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; If your input and references are at different paths than specified, what would you change?
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; If your references are in a different path than what is specified, what would you change?
 
<ul>
 
<ul>
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<div class="mw-collapsible mw-collapsed" style="width:200px">
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<div class="mw-collapsible mw-collapsed" style="width:300px">
 
<li>Answer:</li>
 
<li>Answer:</li>
 
<div class="mw-collapsible-content">
 
<div class="mw-collapsible-content">
 
<ul>
 
<ul>
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<li>You would change <code>IN_DIR</code> & <code>REF_DIR</code> to the new paths</li>
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<li>You would change <code>REF_DIR</code> to the new path</li>
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[[File:gcConf.png|500px]]
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[[File:gcConf.png|800px]]
 
</div>
 
</div>
 
</div>
 
</div>
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Now that we have all of our input files, we need just a simple command to run them
 
Now that we have all of our input files, we need just a simple command to run them
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  ${GC}/gotcloud align --conf ${IN}/gotcloud.conf --numcs 2
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  ${GC}/gotcloud align --conf ${SS}/gotcloud.conf --numcs 2 --base_prefix ${SS} --outdir ${OUT}
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* <code>${GC}/gotcloud</code> runs GotCloud
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* <code>align</code> tells GotCloud you want to run the alignment pipeline.
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* <code>--conf</code> tells GotCloud the name of the configuration file to use.
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** The configuration for this test was downloaded with the seqshop input files.
 
* <code>--numcs</code> means to run 2 samples at a time.
 
* <code>--numcs</code> means to run 2 samples at a time.
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** How many you can run concurrently depends on your system
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** How many you can run concurrently depends on your system.
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* <code>--base_prefix</code> tells GotCloud the prefix to append to relative paths.
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** The Configuration file cannot read environment variables, so we need to tell GotCloud the path to the input files, ${SS}
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** Alternatively, gotcloud.conf could be updated to specify the full paths
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* <code>--out_dir</code> tells GotCloud where to write the output.
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** This could be specified in gotcloud.conf, but to allow you to use the ${OUT} to change the output location, it is specified on the command-line
 +
 
 
[[File:gcalignStart.png|850px]]
 
[[File:gcalignStart.png|850px]]
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[[File:Qplotpdf.png|400px]]
 
[[File:Qplotpdf.png|400px]]
 
<li> Look at the PDF I produced when I ran the whole genome:</li>  
 
<li> Look at the PDF I produced when I ran the whole genome:</li>  
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  evince ${IN}/example/HG00551.wg.qplot.pdf&
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  evince ${SS}/ext/HG00551.wg.qplot.pdf&
 
</ul>
 
</ul>
 
[[File:Qplotpdfwg.png|400px]]
 
[[File:Qplotpdfwg.png|400px]]
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Let's visualize what reads in that area look like using samtools tview:
 
Let's visualize what reads in that area look like using samtools tview:
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  ${GC}/bin/samtools tview ${OUT}/bams/HG00551.recal.bam ${REF}/human.g1k.v37.chr22.fa
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  ${GC}/bin/samtools tview ${OUT}/bams/HG00551.recal.bam ${SS}/ref22/human.g1k.v37.chr22.fa
 
* Type ‘g’  
 
* Type ‘g’  
 
** Type 22:36907000  
 
** Type 22:36907000  
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== Logging Off ==
 
== Logging Off ==
 +
 +
''This section is specifically for the SeqShop Workshop computers.''
 +
<div class="mw-collapsible mw-collapsed" style="width:600px">
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''If you are not running during the SeqShop Workshop, please skip this section.''
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<div class="mw-collapsible-content">
 
To logout of seqshop-server, type:
 
To logout of seqshop-server, type:
 
  exit
 
  exit
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When done, log out of the Windows machine.
 
When done, log out of the Windows machine.
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</div>
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</div>
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== Please provide feedback from Day 1 ==
  −
https://docs.google.com/a/umich.edu/forms/d/1afGq98QToC17CO8k6tXXyybIZFFfNsy8mOX5Tzc2QyM/viewform
 
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